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eBioscience™ BrdU Kit for IHC/ICC Immunofluorescence eFluor™ 570 (Invitrogen™)

This BrdU Kit for immunohistochemistry (IHC)/immunocytochemistry (ICC) with immunofluorescence detection contains the necessary reagents and buffers for identifying and examining proliferating cells by immunohistochemical or immunocytochemical analysis. Cells are labeled in vitro or in vivo with 5-bromo-2’-deoxyuridine (BrdU), a synthetic analog of thymidine, which is incorporated into DNA in place of thymidine during the S-phase of the cell cycle. Following fixation and antigen retrieval steps, cells or tissue sections are stained for BrdU incorporation and visualized using a streptavidin-conjugated fluorophore. This kit has been optimized for IHC with both frozen and formalin-fixed paraffin embedded BrdU-labeled mouse intestine and ICC of BrdU-pulsed HeLa cells grown on culture slides.

Conjugate
eFluo 570

Emit
570 nm

Excite
555 nm

Reported Application
Immunohistochemical Staining of Formalin-Fixed Paraffin Embedded Tissue Sections, Immunohistochemical Staining of Frozen Tissue Sections, Immunocytochemistry, Microscopy

CyQUANT™ XTT Cell Viability Assay (Invitrogen™)

The CyQUANT XTT Cell Viability Assay is a complete, optimized assay that generates a consistent colorimetric detection of viable mammalian cells. The assay kit consists of two reagents, XTT Reagent and Electron Coupling Reagent. XTT Reagent is used to assess cell viability as a function of cellular redox potential, and the electron coupling reagent improves the dynamic range of the assay.

The assay is sufficient for ten 96-well plates and consists of 10 bottles of XTT Reagent and 10 tubes of Electron Coupling Reagent. One bottle of each (sufficient for one 96 well plate) are thawed, mixed together, added to cells, and incubated. The viability signal is detected using an absorbance-based microplate reader. In actively respiring viable mammalian cells, the water-soluble XTT reagent is reduced and converted to an orange-colored formazan product.

More tools for microplate-based detection of viability ›

Features of the CyQUANT XTT Cell Viability Assay include:
• Highly reproducible, absorbance-based viability assay
• Improved dynamic range and sensitivity compared to other colorimetric viability assays
• Simple mix-and-read format, configured for one 96-well plate at a time
• Non-toxic, continuous assay—multiple time points can be collected, no cell lysis

Measuring changes in cell viability is a fundamental method for assessing cell health, determining genotoxicity, and evaluating anti-cancer drugs. In addition to being cost effective, colorimetric viability assays generate consistent results with low well-to-well variability. Unlike other commercially available colorimetric mammalian cell viability assays, XTT-based assays display a high dynamic range and low variability. In addition to improved performance, the assay is a continuous assay and requires no cell lysis or solubilization.

The CyQUANT XTT Cell Viability Assay is a complete and optimized kit for the detection of mammalian cell viability. The assay kit includes XTT Reagent (2,3-bis-(2-methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide) and Electron Coupling Reagent. The XTT reagent, which is a tetrazolium-based compound, is sensitive to cellular redox potential and in the presence of actively respiring cells converts from a water-soluble compound to an orange-colored formazan product. The sensitivity and consistency of the assay is significantly increased when used with the electron coupling reagent.

Since no cell lysis or solubilization are required for detection, the XTT assay can be employed as a continuous assay and several time points can be analyzed. Additionally, the CyQUANT XTT assay generates viability results from various types of cell lines, including primary and suspension cells.

For each 96-well plate, one bottle of XTT Reagent and one tube of Electron Coupling Reagent are thawed, mixed to create a stock solution, and added to cells, with cell viability detected after a 2–4 hour incubation. The assay kit supplies all the materials needed for ten 96-well plates.

It is recommended that the XTT/Electron Coupling reagent stock solution be used soon after mixing. Assay performance is compromised and sensitivity is reduced if the stock solution is kept at room temperature for extended periods of time or subject to cycles of freeze/thaw. To ensure assay performance and reproducibility, the CyQUANT XTT Cell Viability Assay consists of separate XTT and Electron Coupling reagents that are mixed.

eBioscience™ BrdU Kit for IHC/ICC Immunofluorescence eFluor™ 615 (Invitrogen™)

This BrdU Kit for immunohistochemistry (IHC)/immunocytochemistry (ICC) with immunofluorescence detection contains the necessary reagents and buffers for identifying and examining proliferating cells by immunohistochemical or immunocytochemical analysis. Cells are labeled in vitro or in vivo with 5-bromo-2’-deoxyuridine (BrdU), a synthetic analog of thymidine, which is incorporated into DNA in place of thymidine during the S-phase of the cell cycle. Following fixation and antigen retrieval steps, cells or tissue sections are stained for BrdU incorporation and visualized using a streptavidin-conjugated fluorophore. This kit has been optimized for IHC with both frozen and formalin-fixed paraffin embedded BrdU-labeled mouse intestine and ICC of BrdU-pulsed HeLa cells grown on culture slides.

Conjugate
eFluor 615

Emit
615 nm

Excite
595 nm

Reported Application
Immunohistochemical Staining of Formalin-Fixed Paraffin Embedded Tissue Sections, Immunohistochemical Staining of Frozen Tissue Sections, Microscopy, Immunocytochemistry

Arcturus™ Paradise™ PLUS 2 Round, Cy5 labeling (Applied Biosystems™)

The ARCTURUS® Paradise® PLUS Reagent System with Microarray Labeling permits unprecedented gene expression analysis of millions of previously inaccessible samples. The Paradise® PLUS Reagent System is an integrated solution for gene expression studies from formalin-fixed, paraffin-embedded (FFPE) tissues, including reagents for tissue staining, RNA isolation and RNA amplification. Amplify RNA from as little as 5 ng of fixed RNA using methods that have been specially optimized to overcome the challenges of cross-linked templates.

Key product features:
• Microarray Labeling, Gene Expression Profiling
• Flexibility in Assay Format
• Integrated Systems for Microgenomics

Microarray Labeling, Gene Expression Profiling
The Paradise® PLUS RNA Amplification Kit may be configured to include Turbo Labeling™ for use in microarray experiments. Turbo Labeling™ Kits offer a proprietary, non-enzymatic technology optimized for the labeling of unmodified, amplified RNA (aRNA) for gene expression profiling. The unmodified aRNA is labeled post-amplification, thereby avoiding the need to incorporate modified nucleotides during RNA amplification. The use of unmodified nucleotides in the amplification process results in unmodified aRNA with higher yields and longer aRNA fragments, thus providing better representation of the mRNA transcript for downstream analysis.

Flexibility in Assay Format
Turbo Labeling™ Kits include reagents to support labeling of 12 samples using either Cy™3⁄Cy™5 dyes or Biotin for hybridization to cDNA or oligonucleotide arrays.

Integrated Systems for Microgenomics
The Paradise® PLUS Reagent System is validated as part of the ARCTURUS® Complete System for Microgenomics®, an integrated solution for utilizing small quantities of RNA for gene expression analysis, and features the ARCTURUS,sup>XT™ Laser Capture Microdissection (LCM) Instrument to procure pure cell populations. ARCTURUS®' Systems for Microgenomics enable accurate and sensitive microarray assays that reveal molecular otherwise obscured in whole tissue samples or cell mixtures.

For Research Use Only. Not for use in diagnostics procedures.

Escherichia coli (K-12 strain) BioParticles™, BODIPY™ FL conjugate (Invitrogen™)

The Molecular Probes® BioParticles® product line consists of a series of fluorescently labeled, heat- or chemically killed bacteria and yeast in a variety of sizes, shapes, and natural antigenicities. These fluorescent BioParticles® products have been employed to study phagocytosis by fluorescence microscopy, quantitative spectrofluorometry, and flow cytometry.

We offer E. coli (K-12 strain), S. aureus (Wood strain, without protein A) and zymosan (S. cerevisiae) BioParticles® conjugates covalently labeled with a variety of different fluorophores (special care has been taken to remove free dye after conjugation). Unlike the fluorescence of fluorescein-labeled BioParticles® conjugates, which is partially quenched in acidic environments, the fluorescence of the Alexa Fluor®, BODIPY® FL, tetramethylrhodamine and Texas Red® dye conjugates is uniformly intense over the pH range from 3 to 10.

BioParticles Specifications:
• Label (Ex/Em): BODIPY® FL (~505/513 nm)
• Particle: E. coli (K-12 strain)
Opsonizing reagent available


Using BioParticles Products
BioParticles® conjugates are provided as lyophilized powders. There are approximately 3 x 108 E. coli or S. aureus particles per mg solid and approximately 2 x 107 zymosan particles per mg solid. BioParticles® conjugates can be reconstituted in the buffer of your choice for use in phagocytosis assays. The fluorescence of BioParticles® conjugates that are bound to the surface of the cell (but not internalized) can be quenched by ethidium bromide, trypan blue, or other quenchers. In addition to cellular applications, fluorescent BioParticles® conjugates may be effective as flow cytometry calibration references when sorting bacteria and yeast mutants. These small particles may also be useful references for light scattering studies because their sizes and shapes differ in characteristic ways.

Find More BioParticles® Products
We offer a large range of dye-labeled and unlabeled E. coli (K-12 strain), S. aureus (Wood strain, without protein A), and zymosan (S. cerevisiae) BioParticles® products. Find out about these products and their applications by reviewing Probes for Following Receptor Binding and Phagocytosis—Section 16.1 in the Molecular Probes® Handbook.

For pH-sensitive endocytosis assays, see our pHrodo™ BioParticles® conjugates.

For Research Use Only. Not for human or animal therapeutic or diagnostic use.

Arcturus™ Paradise™ PLUS 2 Round Kit, no solvents (Applied Biosystems™)

The ARCTURUS® Paradise® PLUS Reagent System permits unprecedented gene expression analysis of millions of previously inaccessible samples. It is an integrated solution for gene expression studies from formalin-fixed, paraffin-embedded (FFPE) tissues, including reagents for tissue staining, RNA isolation and RNA amplification. Amplify RNA from as little as 5 ng of fixed RNA using methods that have been specially optimized to overcome the challenges of cross-linked templates.

This kit provides reagents for 2-round in-vitro transcription.

Key product features:
• Accelerated discovery process – unlock the tissue archives to access previously inaccessible samples
• Simplified procedure – achieve unprecedented results in six steps
• Streamlined application – integrate seamlessly into current workflow
• Efficient workflow – secure downstream success with the Paradise® QC Kit
• Comprehensive system – benefit from the only complete system for formalin-fixed samples

Unlock the Tissue Archives
Currently, tens of millions of tissue samples with known outcomes are available in tissue archives. The Paradise® PLUS Reagent System enables microarray and qRT-PCR analysis of these previously inaccessible samples, allowing retrospective studies to accelerate discoveries.

Achieve Unprecedented Results in Six Steps
Only six steps are required to take you from sample to results (See Figure 1).

Integrate Seamlessly into Current Workflow
The Paradise® PLUS Reagent System is compatible with all major microarray platforms. Robust and reproducible results are achieved using tools and techniques commonly employed in today’s research laboratories (See Figures 2, 3, 4, and 5).

Secure Downstream Success with the Paradise® QC Kit
Not all FFPE samples contain high quality RNA. Identify the best samples for your studies using a simple, real-time PCR research technique, and then proceed with confidence using these pre-qualified samples (See Figure 6).

Benefit from the Only Complete System for Formalin-fixed Samples
The Paradise® PLUS Reagent System is the only available solution that provides all the components necessary to prepare RNA from formalin-fixed, paraffin-embedded tissue samples for gene expression analysis. To ensure successful microarray results, the system includes reagents optimized for exceptional recovery of RNA and superior RNA amplification.

Integrated Systems for Microgenomics
The Paradise® PLUS Reagent System is a validated as part of the ARCTURUS® Complete System for Microgenomics®, an integrated solution for utilizing small quantities of RNA for gene expression analysis. This system features the ARCTURUSXT™ Laser Capture Microdissection (LCM) Instrument to procure pure cell populations. ARCTURUS® Systems for Microgenomics enable accurate and sensitive microarray assays that reveal molecular signatures otherwise obscured in whole tissue samples or cell mixtures.

For Research Use Only. Not for use in diagnostics procedures.

Membrane Permeability Dead Cell Apoptosis Kit with PO-PRO™-1 and 7-Aminoactinomycin D, for flow cytometry (Invitrogen™)

This flow cytometry product detects apoptosis on the basis of changes that occur in the permeability of cell membranes using PO-PRO™-1 and 7-aminoactinomycin D (7-AAD) nucleic acid stains. After staining a cell population with PO-PRO™-1 dye and 7-AAD, apoptotic cells show violet fluorescence, dead cells show violet and red fluorescence, and live cells show little or no fluorescence. These populations can easily be distinguished by a flow cytometer that uses both a violet laser and the 488 nm line of an argon-ion laser for excitation. Each kit contains sufficient reagents for approximately 200 flow cytometry tests.

View a selection guide for all apoptosis assays for flow cytometry.

Image-iT™ LIVE Green Caspase-3 and -7 Detection Kit, for microscopy (Invitrogen™)

The Image-iT™ LIVE Green Caspase-3 and -7 Detection Kit employs a novel approach to detect active caspases that is based on a fluorescent inhibitor of caspases (FLICA™) methodology, essentially an affinity label. The reagent associates a fluoromethyl ketone (FMK) moiety, which can react covalently with a cysteine, with a caspase-specific amine acid sequence. For caspase-3 and -7, this recognition sequence is aspartic acid-glutamic acid-valine-aspartic acid (DEVD). A fluorescein group is attached as a reporter. The FLICA reagent is thought to interact with the enzymatic reactive center of an activated caspase via the recognition sequence, and then to attach covalently through the FMK moiety. The FLICA inhibitor is cell permeant and noncytotoxic. Unbound FLICA molecules diffuse out of the cell and are washed away; the remaining green-fluorescent signal is a direct measure of the amount of active caspase that was present at the time the inhibitor was added.

eBioscience™ Annexin V Apoptosis Detection Kit PerCP-eFluor™ 710 (Invitrogen™)

Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS) in all mammalian species. Under normal physiologic conditions, PS is predominantly located in the inner leaflet of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution across the phospholipid bilayer and is translocated to the extracellular membrane leaflet marking cells as targets of phagocytosis. Once on the outer surface of the membrane, PS can be detected by fluorescently labeled Annexin V in a calcium-dependent manner.

In early-stage apoptosis, the plasma membrane excludes viability dyes such as propidium iodide (PI), 7-AAD, or Fixable Viability Dyes such as eFluor™ 660 or eFluor™ 780. These cells will stain with Annexin V but not a viability dye, thus distinguishing cells in early apoptosis. However, in late stage apoptosis, the cell membrane loses integrity thereby allowing Annexin V to also access PS in the interior of the cell. A viability dye can be used to resolve these late-stage apoptotic and necrotic cells (Annexin V, viability dye-positive) from the early-stage apoptotic cells (Annexin V positive, viability dye-negative).

Due to the emission spectrum of PerCP-eFluor™ 710, the Annexin V Apoptosis Detection Kit PerCP-eFluor™ 710 is not compatible with propidium iodide and 7-AAD. It is recommended to substitute a Fixable Viability Dye such as eFluor™ 660 or eFluor™ 780 in their place.

Not included:
Fixable Viability Dye eFluor™ 660 (cat. 65-0864)
Fixable Viability Dye eFluor™ 780 (cat. 65-0865)

Note: Fixable Viability Dye eFluor™ 450 is not recommended for use with Annexin V Apoptosis Detection Kits.

Reported Application
Flow Cytometric Analysis

eBioscience™ Annexin V Apoptosis Detection Kit FITC (Invitrogen™)

Annexins are a family of calcium-dependent phospholipid-binding proteins that preferentially bind phosphatidylserine (PS). Under normal physiologic conditions, PS is predominantly located in the inner leaflet of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution across the phospholipid bilayer and is translocated to the extracellular membrane leaflet marking cells as targets of phagocytosis. Once on the outer surface of the membrane, PS can be detected by fluorescently labeled Annexin V in a calcium-dependent manner.

In early-stage apoptosis, the plasma membrane excludes viability dyes such as propidium iodide (PI), 7-AAD, or Fixable Viability Dyes such as eFluor™ 660 or eFluor™ 780. These cells will stain with Annexin V but not a viability dye, thus distinguishing cells in early apoptosis. However, in late stage apoptosis, the cell membrane loses integrity thereby allowing Annexin V to also access PS in the interior of the cell. A viability dye can be used to resolve these late-stage apoptotic and necrotic cells (Annexin V, viability dye-positive) from the early-stage apoptotic cells (Annexin V positive, viability dye-negative).

Note: Fixable Viability Dye eFluor™ 450 is not recommended for use with Annexin V Apoptosis Detection Kits.

Reported Application
Flow Cytometric Analysis

Arcturus™ Paradise™ PLUS 2 Round, Cy5 labeling, no solvents (Applied Biosystems™)

The Arcturus® Paradise® PLUS Reagent System with Microarray Labeling permits unprecedented gene expression analysis of millions of previously inaccessible samples. The Paradise® PLUS Reagent System is an integrated solution for gene expression studies from formalin-fixed, paraffin-embedded (FFPE) tissues, including reagents for tissue staining, RNA isolation and RNA amplification. Amplify RNA from as little as 5 ng of fixed RNA using methods that have been specially optimized to overcome the challenges of cross-linked templates.

Key product features:
• Microarray Labeling, Gene Expression Profiling
• Flexibility in Assay Format
• Integrated Systems for Microgenomics

Microarray Labeling, Gene Expression Profiling
The Paradise® PLUS RNA Amplification Kit may be configured to include Turbo Labeling™ for use in microarray experiments. Turbo Labeling Kits offer a proprietary, non-enzymatic technology optimized for the labeling of unmodified, amplified RNA (aRNA) for gene expression profiling. The unmodified aRNA is labeled post-amplification, thereby avoiding the need to incorporate modified nucleotides during RNA amplification. The use of unmodified nucleotides in the amplification process results in unmodified aRNA with higher yields and longer aRNA fragments, thus providing better representation of the mRNA transcript for downstream analysis.

Flexibility in Assay Format
Turbo Labeling Kits include reagents to support labeling of 12 samples using either Cy™3⁄Cy™5 dyes or Biotin for hybridization to cDNA or oligonucleotide arrays.

Integrated Systems for Microgenomics
The Paradise® PLUS Reagent System is validated as part of the Arcturus® Complete System for Microgenomics®, an integrated solution for utilizing small quantities of RNA for gene expression analysis, and features the ArcturusXT™ Laser Capture Microdissection (LCM) Instrument to procure pure cell populations. Arcturus' Systems for Microgenomics enable accurate and sensitive microarray assays that reveal molecular otherwise obscured in whole tissue samples or cell mixtures.

For Research Use Only. Not for use in diagnostics procedures.

Intracellular pH Calibration Buffer Kit (Invitrogen™)

The Intracellular pH Calibration Buffer Kit enables the quantitation of intracellular pH when used in conjunction with pHrodo™ Red AM, pHrodo™ Green AM, or BCECF intracellular pH indicators. pHrodo™ Red AM and pHrodo™ Green AM are weakly fluorescent at neutral pH but increasingly fluorescent as the pH drops. Modification of pHrodo™ red and green dyes with AM ester groups results in an uncharged molecule that can permeate cell membranes. Once inside the cell, the lipophilic blocking groups are cleaved by nonspecific esterases, resulting in a compound that is retained within the intracellular space. Fluorescence intensity of the probes is then an indicator of intracellular pH. Subsequent use of the Intracellular pH Calibration Buffer Kit allows this intracellular pH to be quantified.

The Intracellular pH Calibration Buffer Kit contains a range of pH calibration buffers (pH 4.5, 5.5, 6.5, and 7.5), as well as valinomycin and nigericin, which help equilibrate the pH inside and outside of cells. After using pHrodo™ Red AM or pHrodo™ Green AM and measuring fluorescence intensity under experimental conditions, the cells are then resuspended in one of the calibration buffers to which valinomycin and nigericin have been added, and the fluorescence intensity measured again. This is repeated with the remaining calibration buffers so that a standard curve can be created and used to pinpoint the intracellular pH associated with the experimental conditions.

LIVE/DEAD™ Fixable Green Dead Cell Stain Kit, for 488 nm excitation (Invitrogen™)

The LIVE/DEAD™ Fixable Green Dead Cell Stain Kit is used to determine the viability of cells prior to the fixation and permeabilization required for intracellular antibody staining or prior to elimination of biohazardous materials using formaldehyde fixation. This kit has been optimized and validated for use with a blue laser flow cytometer.

• Stable—dyes are freeze dried in separate vials to maintain stability

• Robust—staining pattern is the same before and after fixation

• Bright signal—allows for easy distinction between live/dead cells in single channel

View a selection guide for all fixable viability dyes for flow cytometry.

Stable
Unlike products that are sold in solution, the LIVE/DEAD™ Fixable Green Stain has been conveniently packaged in 40-test vials to help ensure the stability and performance of the dye over time. Amine reactive dyes in solution will lose their effectiveness over a short period of time, therefore it is recommended to completely use the vial once rehydrated. If this is not possible, aliquot the vials in small volumes and store at -80°C, avoiding freeze-thaw cycles.

Robust
Dead cell discriminator stains can lose sensitivity after treatment with fixatives such as formaldehyde or ethanol-based fixation methods required for intracellular phosphorylation studies. The LIVE/DEAD™ Fixable Green Stain is an amine reactive dye that binds covalently to intracellular and extracellular amines, and the staining pattern is preserved following formaldehyde fixation.

Optimal brightness
The LIVE/DEAD™ Fixable Green stain was selected based on its fluorescent properties to provide a bright signal when excited with a blue 488 nm laser. The green-fluorescent reactive dye has an excitation maximum of ~495 nm making it ideal for use with the blue laser and an emission of ~520 nm allowing collection in the FITC channel. Since live and dead cells can be discriminated using a single dye and a single channel of a flow cytometer, it is an ideal choice for multi-color experiments.

How it works
In cells with compromised membranes, the dye reacts with free amines both in the cell interior and on the cell surface, yielding intense fluorescent staining. In viable cells, the dye's reactivity is restricted to the cell-surface amines, resulting in less intense fluorescence. The difference in intensity is typically greater than 50-fold between live and dead cells, allowing for easy discrimination.

Colors available
LIVE/DEAD™ Fixable Dead Cell Stains are available in a wide variety of colors to meet your multi-color panel needs.

eBioscience™ Annexin V-Biotin Apoptosis Detection Kit (Invitrogen™)

Annexins are a family of calcium-dependent phospholipid-binding proteins, which bind to phosphatidylserine (PS) to identify apoptotic cells. In healthy cells, PS is predominantly located along the cytosolic side of the plasma membrane. Upon initiation of apoptosis, PS loses its asymmetric distribution in the phospholipid bilayer and translocates to the extracellular membrane, which is detectable with fluorescently labeled Annexin V. In early stages of apoptosis, the plasma membrane excludes viability dyes such as propidium iodide (PI) and 7-AAD, therefore cells which display only Annexin V staining (PI/7-AAD negative) are in early stages of apoptosis. During late-stage apoptosis, loss of cell membrane integrity allows Annexin V binding to cytosolic PS, as well as cell uptake of PI and 7-AAD. Annexin V staining, paired with 7-AAD or PI is widely used to identify apoptotic stages by flow cytometry.

Host
E.coli

Conjugate
Biotin

Purity
> 98% pure

Molecular Mass
35.8 kDa

Reported Application
Flow Cytometric Analysis

2-NBDG (2-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)Amino)-2-Deoxyglucose) (Invitrogen™)

2-NBDG is a fluorescent glucose analog that has been used to monitor glucose uptake in live cells, as an indicator of cell viability. Although sensitive to its environment NBD fluorescence typically displays excitation/emission maxima of ~465/540 nm and can be visualized using optical filters designed for fluorescein.